Synthesis of chromogenic probes for detection and assay of diglycosidases

Matej Cvečko 1 Peter Kis 1 Mária Mastihubová 1 Vladimír Mastihuba 1

1Chemický ústav SAV, Bratislava, Slovenská republika

Diglycosidases represent a special group of glycoside hydrolases characterized by their unique way of hydrolysis of natural products, such as rutin, hesperidin, furcatin, vicianin, etc.[1] Four yet known diglycosidases, namely rutinosidase/hesperidinase, acuminosidase, vicianosidase, and primeverosidase may be considered as endo-β-glucosidases, [2] since they hydrolyze β-glycosidic bond between aglycone and glucose, with another saccharide moiety (e.g. rhamnopyranosyl, apiofuranosyl, xylopyranosyl, arabinopyranosyl) [3] bound in the position 6-O of the glucopyranoside ring.

In this work we synthesized two derivatives of natural disaccharides 4-nitrophenyl β-acuminoside (1) and 2-nitrophenyl β-robinobioside (2) as chromogenic probes of two diglycosidases – acuminosidase and robinobiosidase, respectively. The key steps of synthesis were glycosylations of appropriate peracetylated nitrophenyl glycosides previously deprotected in position 6, with peracetylated D-apiofuranosyl donor or D-rhamnopyranosyl donor, respectively. Two different glycosylation methods were used, resulting in good yields and excellent stereoselectivity. After deprotection of glycosylation products we obtained 43 % of 1 and 83% of 2 overall in two steps. We intend to use these compounds as chromogenic probes for two diglycosidases – acuminosidase and robinobiosidase.

This work was supported by the Slovak Research and Development Agency under the contract No. APVV-18-0188 and by the Slovak Grant Agency for Science VEGA (grant number 2/0126/19). The work was inspired by scientific interactions that evoved within the COST Action CA18103 - Innovation with Glycans: new frontiers from synthesis to new biological targets (INNOGLY).
[1] Sakata, K.; Mizutani, M.; Ma, S. J.; Hiratake, J. Methods Enzymol., 2003, 363, 444-459.
[2] Mazzaferro, L. S.; Breccia, J. D. Biocatal. Biotrans. 2011, 29, 103-112.
[3] Koseki, T.; Ishikawa, M.; Kawasaki, M.; Shiono, Y. Appl. Microbiol. Biotechnol. 2018, 102, 8717-8723.